Novel anti-inflammatory androstane derivative

ABSTRACT

According to one aspect of the invention, there is provided a pharmaceutical formulation for administration by inhalation comprising a compound of formula (I),  
                 
 
     or a solvate thereof, together with a long-acting β 2 -adrenoreceptor agonist which formulation has a therapeutically useful effect in the treatment of inflammatory disorders of the respiratory tract over a period of 24 hours or more.

[0001] This application is a Continuation-in-part of U.S. patentapplication Ser. No. 09/958050 filed on Oct. 2, 2001, which is basedupon International Patent Application No. PCT.GB01.03495 filed Aug. 3,2001, which claims priority to United Kingdom Patent Application No. GB0019172.6 filed Aug. 5, 2000.

[0002] The present invention relates to a novel anti-inflammatory andanti-allergic compound of the androstane series and to processes for itspreparation. The present invention also relates to pharmaceuticalformulations containing the compound and to therapeutic uses thereof,particularly for the treatment of inflammatory and allergic conditions.

[0003] Glucocorticoids which have anti-inflammatory properties are knownand are widely used for the treatment of inflammatory disorders ordiseases such as asthma and rhinitis. For example, U.S. Pat. No.4,335,121 discloses 6α,9α-Difluoro-17α-(1-oxopropoxy)-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (known by the generic name of fluticasonepropionate) and derivatives thereof. The use of glucocorticoidsgenerally, and especially in children, has been limited in some quartersby concerns over potential side effects. The side effects that arefeared with glucocorticoids include suppression of theHypothalamic-Pituitary-Adrenal (HPA) axis, effects on bone growth inchildren and on bone density in the elderly, ocular complications(cataract formation and glaucoma) and skin atrophy. Certainglucocorticoid compounds also have complex paths of metabolism whereinthe production of active metabolites may make the pharmacodynamics andpharmacokinetics of such compounds difficult to understand. Whilst themodern steroids are very much safer than those originally introduced, itremains an object of research to produce new molecules which haveexcellent anti-inflammatory properties, with predictable pharmacokineticand pharmacodynamic properties, with an attractive side effect profile,and with a convenient treatment regime.

[0004] We have now identified a novel glucocorticoid compound whichsubstantially meets these objectives.

[0005] Thus, according to one aspect of the invention, there is provideda compound of formula (I)

[0006] and solvates thereof.

[0007] The chemical name of the compound of formula (I) is 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester.

[0008] References hereinafter to the compound according to the inventioninclude both the compound of formula (I) and solvates thereof,particularly pharmaceutically acceptable solvates.

[0009] The compound of formula (I) has potentially beneficialanti-inflammatory or anti-allergic effects, particularly upon topicaladministration, demonstrated by, for example, its ability to bind to theglucocorticoid receptor and to illicit a response via that receptor,with long acting effect. Hence, the compound of formula (I) is useful inthe treatment of inflammatory and/or allergic disorders, especially inonce-per-day therapy.

[0010] Compound (I) undergoes highly efficient hepatic metabolism toyield the 17-βcarboxylic acid (X) as the sole major metabolite in ratand human in vitro systems. This metabolite has been synthesised anddemonstrated to be >1000 fold less active than the parent compound in invitro functional glucocorticoid assays.

[0011] This efficient hepatic metabolism is reflected by in vivo data inthe rat, which have demonstrated plasma clearance at a rate approachinghepatic blood flow and an oral bioavailability of <1%, consistent withextensive first-pass metabolism.

[0012] In vitro metabolism studies in human hepatocytes havedemonstrated that compound (I) is metabolised in an identical manner tofluticasone propionate but that conversion of (I) to the inactive acidmetabolite occurs approximately 5-fold more rapidly than withfluticasone propionate. This very efficient hepatic inactivation wouldbe expected to minimise systemic exposure in man leading to an improvedsafety profile.

[0013] Inhaled steroids are also absorbed through the lung and thisroute of absorption makes a significant contribution to systemicexposure. Reduced lung absorption could therefore provide an improvedsafety profile. Studies with compound of formula (I) have shownsignificantly lower exposure to compound of formula (I) than withfluticasone propionate after dry powder delivery to the lungs ofanaesthetised pigs.

[0014] An improved safety profile is believed to allow the compound offormula (I) to demonstrate the desired anti-inflammatory effects whenadministered once-per day. Once-per-day dosing is considered to besignificantly more convenient to patients than the twice-per day dosingregime that is normally employed for fluticasone propionate.

[0015] Examples of disease states in which the compound of the inventionhas utility include skin diseases such as eczema, psoriasis, allergicdermatitis, neurodermatitis, pruritis and hypersensitivity reactions;inflammatory conditions of the nose, throat or lungs such as asthma(including allergen-induced asthmatic reactions), rhinitis (includinghayfever), nasal polyps, chronic obstructive pulmonary disease (COPD),interstitial lung disease, and fibrosis; inflammatory bowel conditionssuch as ulcerative colitis and Crohn's disease; and auto-immune diseasessuch as rheumatoid arthritis.

[0016] The compound of the invention may also have use in the treatmentof conjunctiva and conjunctivitis.

[0017] The compound of formula (I) and solvates thereof is expected tobe most useful in the treatment of inflammatory disorders of therespiratory tract eg asthma and COPD, particularly asthma.

[0018] It will be appreciated by those skilled in the art that referenceherein to treatment extends to prophylaxis as well as the treatment ofestablished conditions.

[0019] As mentioned above, the compound of formula (I) is useful inhuman or veterinary medicine, in particular as an anti-inflammatory andanti-allergic agent.

[0020] There is thus provided as a further aspect of the invention thecompound of formula (I) or a physiologically acceptable solvate thereoffor use in human or veterinary medicine, particularly in the treatmentof patients with inflammatory and/or allergic conditions, especially fortreatment once-per-day.

[0021] According to another aspect of the invention, there is providedthe use of the compound of formula (I) or physiologically acceptablesolvate thereof for the manufacture of a medicament for the treatment ofpatients with inflammatory and/or allergic conditions, especially fortreatment once-per-day.

[0022] In a further or alternative aspect, there is provided a methodfor the treatment of a human or animal subject with an inflammatoryand/or allergic condition, which method comprises administering to saidhuman or animal subject an effective amount of the compound of formula(I) or physiologically acceptable solvate thereof, especially foradministration once-per-day.

[0023] The compound according to the invention may be formulated foradministration in any convenient way, and the invention therefore alsoincludes within its scope pharmaceutical compositions comprising thecompound of formula (I) or a physiologically acceptable solvate thereoftogether, if desirable, in admixture with one or more physiologicallyacceptable diluents or carriers. Pharmaceutical compositions suitablefor once-per-day administration are of particular interest.

[0024] Further, there is provided a process for the preparation of suchpharmaceutical compositions which comprises mixing the ingredients.

[0025] The compound according to the invention may, for example, beformulated for oral, buccal, sublingual, parenteral, local or rectaladministration, especially local administration.

[0026] Local administration as used herein, includes administration byinsufflation and inhalation. Examples of various types of preparationfor local administration include ointments, lotions, creams, gels,foams, preparations for delivery by transdermal patches, powders,sprays, aerosols, capsules or cartridges for use in an inhaler orinsufflator or drops (eg eye or nose drops), solutions/suspensions fornebulisation, suppositories, pessaries, retention enemas and chewable orsuckable tablets or pellets (eg for the treatment of aphthous ulcers) orliposome or microencapsulation preparations.

[0027] Advantageously compositions for topical administration to thelung include dry powder compositions and spray compositions.

[0028] Dry powder compositions for topical delivery to the lung byinhalation may, for example, be presented in capsules and cartridges foruse in an inhaler or insufflator of, for example, gelatine. Formulationsgenerally contain a powder mix for inhalation of the compound of theinvention and a suitable powder base (carrier substance) such as lactoseor starch. Use of lactose is preferred. Each capsule or cartridge maygenerally contain between 20 μg-10 μg of the compound of formula (I)optionally in combination with another therapeutically activeingredient. Alternatively, the compound of the invention may bepresented without excipients. Packaging of the formulation may besuitable for unit dose or multi-dose delivery. In the case of multi-dosedelivery, the formulation can be pre-metered (eg as in Diskus, see GB2242134 or Diskhaler, see GB 2178965, 2129691 and 2169265) or metered inuse (eg as in Turbuhaler, see EP 69715). An example of a unit-dosedevice is Rotahaler (see GB 2064336). The Diskus inhalation devicecomprises an elongate strip formed from a base sheet having a pluralityof recesses spaced along its length and a lid sheet hermetically butpeelably sealed thereto to define a plurality of containers, eachcontainer having therein an inhalable formulation containing a compoundof formula (I) optionally in combination with another therapeuticallyactive ingredient preferably combined with lactose. Preferably, thestrip is sufficiently flexible to be wound into a roll. The lid sheetand base sheet will preferably have leading end portions which are notsealed to one another and at least one of the said leading end portionsis constructed to be attached to a winding means. Also, preferably thehermetic seal between the base and lid sheets extends over their wholewidth. The lid sheet may preferably be peeled from the base sheet in alongitudinal direction from a first end of the said base sheet.

[0029] Spray compositions for topical delivery to the lung by inhalationmay for example be formulated as aqueous solutions or suspensions or asaerosols delivered from pressurised packs, such as a metered doseinhaler, with the use of a suitable liquefied propellant. Aerosolcompositions suitable for inhalation can be either a suspension or asolution and generally contain the compound of formula (I) optionally incombination with another therapeutically active ingredient and asuitable propellant such as a fluorocarbon or hydrogen-containingchlorofluorocarbon or mixtures thereof, particularly hydrofluoroalkanes,especially 1,1,1,2-tetrafluoroethane,1,1,1,2,3,3,3-heptafluoro-n-propane or a mixture thereof. The aerosolcomposition may optionally contain additional formulation excipientswell known in the art such as surfactants eg oleic acid or lecithin andcosolvents eg ethanol. One example formulation is excipient free andconsists essentially of (eg consists of) compound of formula (I)(preferably in unsolvated form eg as Form 1) (optionally together with afurther active ingredient) and a propellant selected from1,1,1,2-tetrafluoroethane, 1,1,1,2,3,3,3-heptafluoro-n-propane andmixture thereof. Another example formulation comprises particulatecompound of formula (I), a propellant selected from1,1,1,2-tetrafluoroethane, 1,1,1,2,3,3,3-heptafluoro-n-propane andmixture thereof and a suspending agent which is soluble in thepropellant eg an oligolactic acid or derivative thereof as described inWO94/21229. The preferred propellant is 1,1,1,2-tetrafluoroethane. Asnoted elsewhere in this specification, compound of formula (I) does notappear to form a solvate with 1,1,1,2-tetrafluoroethane. Pressurisedformulations will generally be retained in a canister (eg an aluminiumcanister) closed with a valve (eg a metering valve) and fitted into anactuator provided with a mouthpiece.

[0030] Pressurised aerosol formulations preferably do not compriseparticulate medicament, a propellant and a stabiliser comprising a wateraddition (i.e. water added in addition to nascent formulation water).Pressurised aerosol formulations also preferably do not compriseparticulate medicament, a propellant and a stabiliser comprising anamino acid, a derivative thereof or a mixture thereof.

[0031] Medicaments for administration by inhalation desirably have acontrolled particle size. The optimum particle size for inhalation intothe bronchial system is usually 1-10 μm, preferably 2-5 μm. Particleshaving a size above 20 μm are generally too large when inhaled to reachthe small airways. To achieve these particle sizes the particles ofcompound of formula (I) (and any further therapeutically activeingredient) as produced may be size reduced by conventional means eg bymicronisation. The desired fraction may be separated out by airclassification or sieving. Preferably, the particles will becrystalline, prepared for example by a process which comprises mixing ina continuous flow cell in the presence of ultrasonic radiation a flowingsolution of compound of formula (I) as medicament in a liquid solventwith a flowing liquid antisolvent for said medicament (eg as describedin International Patent Application PCT/GB99/04368) or else by a processwhich comprises admitting a stream of solution of the substance in aliquid solvent and a stream of liquid antisolvent for said substancetangentially into a cylindrical mixing chamber having an axial outletport such that said streams are thereby intimately mixed throughformation of a vortex and precipitation of crystalline particles of thesubstance is thereby caused (eg as described in International PatentApplication PCT/GB00/04327). When an excipient such as lactose isemployed, generally, the particle size of the excipient will be muchgreater than the inhaled medicament within the present invention. Whenthe excipient is lactose it will typically be present as milled lactose,wherein not more than 85% of lactose particles will have a MMD of 60-90μm and not less than 15% will have a MMD of less than 15 μm.

[0032] Formulations for administration topically to the nose (eg for thetreatment of rhinitis) include pressurised aerosol formulations andaqueous formulations administered to the nose by pressurised pump.Formulations which are non-pressurised and adapted to be administeredtopically to the nasal cavity are of particular interest. Theformulation preferably contains water as the diluent or carrier for thispurpose. Aqueous formulations for administration to the lung or nose maybe provided with conventional excipients such as buffering agents,tonicity modifying agents and the like. Aqueous formulations may also beadministered to the nose by nebulisation.

[0033] Other possible presentations include the following:

[0034] Ointments, creams and gels, may, for example, be formulated withan aqueous or oily base with the addition of suitable thickening and/orgelling agent and/or solvents. Such bases may thus, for example, includewater and/or an oil such as liquid paraffin or a vegetable oil such asarachis oil or castor oil, or a solvent such as polyethylene glycol.Thickening agents and gelling agents which may be used according to thenature of the base include soft paraffin, aluminium stearate,cetostearyl alcohol, polyethylene glycols, woolfat, beeswax,carboxypolymethylene and cellulose derivatives, and/or glycerylmonostearate and/or non-ionic emulsifying agents.

[0035] Lotions may be formulated with an aqueous or oily base and willin general also contain one or more emulsifying agents, stabilisingagents, dispersing agents, suspending agents or thickening agents.

[0036] Powders for external application may be formed with the aid ofany suitable powder base (carrier substance), for example, talc, lactoseor starch. Drops may be formulated with an aqueous or non-aqueous basealso comprising one or more dispersing agents, solubilising agents,suspending agents or preservatives.

[0037] If appropriate, the formulations of the invention may be bufferedby the addition of suitable buffering agents.

[0038] The proportion of the active compound of formula (I) in the localcompositions according to the invention depends on the precise type offormulation to be prepared but will generally be within the range offrom 0.001 to 10% by weight. Generally, however for most types ofpreparations advantageously the proportion used will be within the rangeof from 0.005 to 1% and preferably 0.01 to 0.5%. However, in powders forinhalation or insufflation the proportion used will usually be withinthe range of from 0.1 to 5%.

[0039] Aerosol formulations are preferably arranged so that each metereddose or “puff” of aerosol contains 1 μg-2000 μg eg 20 μg-2000 μg,preferably about 20 μg-500 μg of a compound of formula (I) optionally incombination with another therapeutically active ingredient.Administration may be once daily or several times daily, for example 2,3, 4 or 8 times, giving for example 1, 2 or 3 doses each time.Preferably the compound of formula (I) is delivered once or twice daily,more preferably once-per-day. The overall daily dose with an aerosolwill typically be within the range 10 μg-10 mg eg 100 μg-10 mgpreferably, 200 μg-2000 μg.

[0040] Since the compound of formula (I) is long-acting, preferably thecompound will be delivered once-per-day and the dose will be selected sothat the compound has a therapeutic effect in the treatment ofrespiratory disorders (eg asthma or COPD, particularly asthma) over 24hours or more.

[0041] Topical preparations may be administered by one or moreapplications per day to the affected area; over skin areas occlusivedressings may advantageously be used. Continuous or prolonged deliverymay be achieved by an adhesive reservoir system.

[0042] For internal administration the compound according to theinvention may, for example, be formulated in conventional manner fororal, parenteral or rectal administration. Formulations for oraladministration include syrups, elixirs, powders, granules, tablets andcapsules which typically contain conventional excipients such as bindingagents, fillers, lubricants, disintegrants, wetting agents, suspendingagents, emulsifying agents, preservatives, buffer salts, flavouring,colouring and/or sweetening agents as appropriate. Dosage unit formsare, however, preferred as described below.

[0043] Preferred forms of preparation for internal administration aredosage unit forms i.e. tablets and capsules. Such dosage unit formscontain from 0.1 mg to 20 mg preferably from 2.5 to 10 mg of thecompound of the invention.

[0044] The compound according to the invention may in general may begiven by internal administration in cases where systemic adreno-corticaltherapy is indicated.

[0045] In general terms preparations, for internal administration maycontain from 0.05 to 10% of the active ingredient dependent upon thetype of preparation involved. The daily dose may vary from 0.1 mg to 60mg, eg 5-30 mg, dependent on the condition being treated, and theduration of treatment desired.

[0046] Slow release or enteric coated formulations may be advantageous,particularly for the treatment of inflammatory bowel disorders.

[0047] The pharmaceutical compositions according to the invention mayalso be used in combination with another therapeutically active agent,for example, a β₂ adrenoreceptor agonist, an anti-histamine or ananti-allergic. The invention thus provides, in a further aspect, acombination comprising the compound of formula (I) or a physiologicallyacceptable solvate thereof together with another therapeutically activeagent, for example, a β₂-adrenoreceptor agonist, an anti-histamine or ananti-allergic.

[0048] Examples of β₂-adrenoreceptor agonists include salmeterol (eg asracemate or a single enantiomer such as the R-enantiomer), salbutamol,formoterol, salmefamol, fenoterol or terbutaline and salts thereof, forexample the xinafoate salt of salmeterol, the sulphate salt or free baseof salbutamol or the fumarate salt of formoterol. Pharmaceuticalcompositions employing combinations with long-acting β₂-adrenoreceptoragonists (eg salmeterol and salts thereof) are particularly preferred,especially those which have a therapeutic effect (eg in the treatment ofasthma or COPD, particularly asthma) over 24 hours or more.

[0049] Since the compound of formula (I) is long-acting, preferably thecomposition comprising the compound of formula (I) and the long-actingβ₂-adrenoreceptor agonists will be delivered once-per-day and the doseof each will be selected so that the composition has a therapeuticeffect in the treatment of respiratory disorders effect (eg in thetreatment of asthma or COPD, particularly asthma) over 24 hours or more.

[0050] Examples of anti-histamines include methapyrilene or loratadine.

[0051] Other suitable combinations include, for example, otheranti-inflammatory agents eg NSAIDs (eg sodium cromoglycate, nedocromilsodium, PDE4 inhibitors, leukotriene antagonists, iNOS inhibitors,tryptase and elastase inhibitors, beta-2 integrin antagonists andadenosine 2a agonists)) or antiinfective agents (eg antibiotics,antivirals).

[0052] Also of particular interest is use of the compound of formula (I)in combination with a phosphodiesterase 4 (PDE4) inhibitor. ThePDE4-specific inhibitor useful in this aspect of the invention may beany compound that is known to inhibit the PDE4 enzyme or which isdiscovered to act as a PDE4 inhibitor, and which are only PDE4inhibitors, not compounds which inhibit other members of the PDE familyas well as PDE4. Generally it is preferred to use a PDE4 inhibitor whichhas an IC₅₀ ratio of about 0.1 or greater as regards the IC₅₀ for thePDE4 catalytic form which binds rolipram with a high affinity divided bythe IC₅₀ for the form which binds rolipram with a low affinity. For thepurposes of this disclosure, the cAMP catalytic site which binds R and Srolipram with a low affinity is denominated the “low affinity” bindingsite (LPDE 4) and the other form of this catalytic site which bindsrolipram with a high affinity is denominated the “high affinity” bindingsite (HPDE 4). This term “HPDE4” should not be confused with the term“hPDE4” which is used to denote human PDE4. Initial experiments wereconducted to establish and validate a [³H]-rolipram binding assay.Details of this work are given in the Binding Assays described in detailbelow.

[0053] The preferred PDE4 inhibitors of use in this invention will bethose compounds which have a salutary therapeutic ratio, i.e., compoundswhich preferentially inhibit cAMP catalytic activity where the enzyme isin the form that binds rolipram with a low affinity, thereby reducingthe side effects which apparently are linked to inhibiting the formwhich binds rolipram with a high affinity. Another way to state this isthat the preferred compounds will have an IC₅₀ ratio of about 0.1 orgreater as regards the IC₅₀ for the PDE4 catalytic form which bindsrolipram with a high affinity divided by the IC₅₀ for the form whichbinds rolipram with a low affinity. A further refinement of thisstandard is that of one wherein the PDE4 inhibitor has an IC₅₀ ratio ofabout 0.1 or greater; said ratio is the ratio of the IC₅₀ value forcompeting with the binding of 1nM of [³H]R-rolipram to a form of PDE4which binds rolipram with a high affinity over the IC₅₀ value forinhibiting the PDE4 catalytic activity of a form which binds rolipramwith a low affinity using 1 μM[³H]-cAMP as the substrate.

[0054] Examples of useful PDE4 inhibitors are:

[0055](R)-(+)-1-(4-bromobenzyl)-4-[(3-cyclopentyloxy)-4-methoxyphenyl]-2-pyrrolidone;

[0056](R)-(+)-1-(4-bromobenzyl)-4-[(3-cyclopentyloxy)-4-methoxyphenyl]-2-pyrrolidone;

[0057]3-(cyclopentyloxy-4-methoxyphenyl)-1-(4-N′-[N2-cyano-S-methyl-isothioureido]benzyl)-2-pyrrolidone;

[0058] cis4-cyano-4-(3-cyclopentyloxy-4-methoxyphenyl)cyclohexan-1-carboxylicacid];

[0059]cis-[4-cyano-4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-1-ol];

[0060] (R)-(+)-ethyl[4-(3-cyclopentyloxy-4-methoxyphenyl)pyrrolidine-2-ylidene]acetate; and

[0061] (S)-(−)-ethyl[4-(3-cyclopentyloxy-4-methoxyphenyl)pyrrolidine-2-ylidene]acetate.

[0062] Most preferred are those PDE4 inhibitors which have an IC₅₀ ratioof greater than 0.5, and particularly those compounds having a ratio ofgreater than 1.0. Preferred compounds are cis4-cyano-4-(3-cyclopentyloxy-4-methoxyphenyl)cyclohexan-1-carboxylicacid,2-carbomethoxy-4-cyano-4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-1-oneandcis-[4-cyano-4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-1-ol];these are examples of compounds which bind preferentially to the lowaffinity binding site and which have an IC₅₀ ratio of 0.1 or greater.

[0063] Other compounds of interest include:

[0064] Compounds set out in U.S. Pat. No. 5,552,438 issued Sep. 3, 1996;this patent and the compounds it discloses are incorporated herein infull by reference. The compound of particular interest, which isdisclosed in U.S. Pat. No. 5,552,438, iscis-4-cyano-4-[3-(cyclopentyloxy)-4-methoxyphenyl]cyclohexane-1-carboxylicacid (also known as cilomalast) and its salts, esters, pro-drugs orphysical forms; AWD-12-281 from Astra (Hofgen, N. et al. 15th EFMC IntSymp Med Chem (September 6-10, Edinburgh) 1998, Abst P.98); a9-benzyladenine derivative nominated NCS-613 (INSERM); D-4418 fromChiroscience and Schering-Plough; a benzodiazepine PDE4 inhibitoridentified as Cl-1018 (PD-168787; Parke-Davis/Warner-Lambert); abenzodioxole derivative Kyowa Hakko disclosed in WO 9916766; V-11294Afrom Napp (Landells, L. J. et al. Eur Resp J [Annu Cong Eur Resp Soc(September 19-23, Geneva) 1998] 1998, 12(Suppl. 28): Abst P2393);roflumilast (CAS reference No 162401-32-3) and a pthalazinone (WO9947505) from Byk-Gulden; or a compound identified as T-440 (TanabeSeiyaku; Fuji, K. et al. J Pharmacol Exp Ther, 1998, 284(1): 162).

[0065] Phosphodiesterase and Rolipram Binding Assays

[0066] Assay Method 1A

[0067] Isolated human monocyte PDE4 and hrPDE (human recombinant PDE4)was determined to exist primarily in the low affinity form. Hence, theactivity of test compounds against the low affinity form of PDE4 can beassessed using standard assays for PDE4 catalytic activity employing 1μM [³H]cAMP as a substrate (Torphy et al., J. of Biol. Chem., Vol. 267,No. 3 pp 1798-1804, 1992). Rat brain high speed supernatants were usedas a source of protein and both enantiomers of [³H]-rolipram wereprepared to a specific activity of 25.6 Ci/mmol. Standard assayconditions were modified from the published procedure to be identical tothe PDE assay conditions, except for the last of the cAMP: 50 mM TrisHCl (pH 7.5), 5 mM MgCl₂, 50 μM 5′-AMP and 1 nM of [³H]-rolipram (Torphyet al., J. of Biol. Chem., Vol. 267, No. 3 pp 1798-1804, 1992). Theassay was run for 1 hour at 30° C. The reaction was terminated and boundligand was separated from free ligand using a Brandel cell harvester.Competition for the high affinity binding site was assessed underconditions that were identical to those used for measuring low affinityPDE activity, expect that [³H]-cAMP was not present.

[0068] Assay Method 1B

[0069] Measurement of Phosphodiesterase Activity

[0070] PDE activity was assayed using a [³H]cAMP SPA or [³H]cGMP SPAenzyme assay as described by the supplier (Amersham Life Sciences). Thereactions were conducted in 96-well plates at room temperature, in 0.1ml of reaction buffer containing (final concentrations): 50 mM Tris-HCl,pH 7.5, 8.3 mM MgCl₂, 1.7 mM EGTA, [³H]cAMP or [³H] cGMP (approximately2000 dpm/pmol), enzyme and various concentrations of the inhibitors. Theassay was allowed to proceed for 1 hr and was terminated by adding 50 μlof SPA yttrium silicate beads in the presence of zinc sulfate. Theplates were shaken and allowed to stand at room temperature for 20 min.Radiolabeled product formation was assessed by scintillationspectrometry.

[0071] [³H]R-Rolipram Binding Assay

[0072] The [³H]R-rolipram binding assay was performed by modification ofthe method of Schneider and co-workers, see Nicholson, et al., TrendsPharmacol. Sci., Vol. 12, pp. 19-27 (1991) and McHale et al., Mol.Pharmacol., Vol. 39, 109-113 (1991). R-Rolipram binds to the catalyticsite of PDE4 see Torphy et al., Mol. Pharmacol., Vol. 39, pp. 376-384(1991). Consequently, competition for [³H]R-rolipram binding provides anindependent confirmation of the PDE4 inhibitor potencies of unlabeledcompetitors. The assay was performed at 30° C. for 1 hr in 0.5 μl buffercontaining (final concentrations): 50 mM Tris-HCl, pH 7.5, 5 mM MgCl₂,0.05% bovine serum albumin, 2 nM [³H]R-rolipram (5.7×104 dpm/pmol) andvarious concentrations of non-radiolabeled inhibitors. The reaction wasstopped by the addition of 2.5 ml of ice-cold reaction buffer (without[³H]-R-rolipram) and rapid vacuum filtration (Brandel Cell Harvester)through Whatman GF/B filters that had been soaked in 0.3%polyethylenimine. The filters were washed with an additional 7.5 ml ofcold buffer, dried, and counted via liquid scintillation spectrometry.

[0073] The invention thus provides, in a further aspect, a combinationcomprising the compound of formula (I) or a physiologically acceptablesolvate thereof together with a PDE4 inhibitor.

[0074] The combination referred to above may conveniently be presentedfor use in the form of a pharmaceutical formulation and thuspharmaceutical formulations comprising a combination as defined abovetogether with a physiologically acceptable diluent or carrier representa further aspect of the invention.

[0075] The compound according to the invention in combination withanother therapeutically active ingredient as described above may beformulated for administration in any convenient way, and the inventiontherefore also includes within its scope pharmaceutical compositionscomprising the compound of formula (I) or a physiologically acceptablesolvate thereof in combination with another therapeutically activeingredient together, if desirable, in admixture with one or morephysiologically acceptable diluents or carriers. The preferred route ofadministration for inflammatory disorders of the respiratory tract willgenerally be administration by inhalation.

[0076] Further, there is provided a process for the preparation of suchpharmaceutical compositions which comprises mixing the ingredients.

[0077] The individual compounds of such combinations may be administeredeither sequentially in separate pharmaceutical compositions as well assimultaneously in combined pharmaceutical formulations. Appropriatedoses of known therapeutic agents will be readily appreciated by thoseskilled in the art.

[0078] Surprisingly, the compound of formula (I) has demonstrated asignificant propensity to form solvates with commonly used organicsolvents. Such solvates are essentially stoichiometric eg the ratio ofcompound of formula (I) to solvent is close to 1:1 eg according toApplicant's analysis has been determined to be in the range 0.95-1.05:1.For example, we have prepared solvates with solvents such as acetone,dimethylformamide (DMF), dimethylacetamide (DMAc), tetrahydrofuran(THF), N-methyl-2-pyrrolidone, isopropanol and methylethylketone. Thesolvation of compound of formula (I) is not predictable however since wehave found that even though it does form a solvate with isopropanol itdoes not appear to form a solvate with ethanol or methanol. Furthermoreit does not appear to form a solvate with 1,1,1,2-tetrafluoroethane,ethylacetate, methylacetate, toluene, methylisobutylketone (MIBK) orwater either. However due to the toxicity of many organic solvents ithas been necessary to develop special final stage processing conditions(discussed later) in order to permit the compound of formula (I) to beproduced in unsolvated form. Thus according to another aspect of theinvention there is provided a compound of formula (I) in unsolvatedform.

[0079] Surprisingly we have also discovered that the compound of formula(I) in unsolvated form may exist in a number of polymorphic forms.Specifically we have identified polymorphic forms which may bedistinguished by means of X-Ray Powder Diffraction (XRPD) which we havenamed as Form 1, Form 2 and Form 3. Form 3 appears to be an unstableminor polymorphic modification of Form 2. Broadly speaking the Forms arecharacterised in their XRPD profiles as follows:

[0080] Form 1: Peak at around 18.9 degrees 2Theta

[0081] Form 2: Peaks at around 18.4 and 21.5 degrees 2Theta.

[0082] Form 3: Peaks at around 18.6 and 19.2 degrees 2Theta.

[0083] Within the range 21-23 degrees 2Theta Form 3 shows a single peakwhereas Form 2 shows a pair of peaks. A peak at 7 degrees 2Theta ispresent in all cases however it is present at much higher intensity inthe case of Forms 2 and 3 than is the case for Form 1.

[0084] The XRPD patterns of the polymorphs are shown overlaid in FIG. 1.The conversion of Form 2 to Form 1 with time in an aqueous slurry atambient temperature is shown in FIG. 2. In the conversion of Form 2 toForm 1 the loss of a peak characteristic of Form 2 (labelled B) ataround 18.4 degrees 2Theta, a marked reduction in intensity in the peakat around 7 degrees 2Theta (labelled A) and the appearance of a peakcharacteristic of Form 1 (labelled C) at around 18.9 degrees 2Theta areparticularly noticeable.

[0085] The temperature dependence of Form 3 is shown in FIG. 4. Thetemperature was varied according to the profile shown in FIG. 5. FromFIG. 4 it can be seen that Form 3 converts first to Form 2 over thetemperature range 30-170° C. and then converts to Form 1 over thetemperature range 170-230° C. In the conversion of Form 3 to Form 2 thedivision of one peak in the range 21-23 degrees 2Theta into two peakswithin the same range and the shifting leftwards of the peak at around18.6 degrees 2Theta to around 18.4 degrees 2Theta are particularlynoticeable. In the conversion of Form 2 to Form 1 similar changes tothose noted in the previous paragraph may be observed.

[0086] The differential scanning calorimetry (DSC) and thermalgravimetric analysis (TGA) profiles of Form 1 are shown in FIG. 3. Theprofiles are characterised by a transition at around 280-300° C.(typically close to 298° C.) corresponding to an endothermic event inthe DSC and chemical degradation in the TGA. The DSC profiles of Forms 2and 3 were not materially different under the conditions of theexperiments performed and thus DSC is not a suitable technique fordistinguishing between the 3 Forms. In FIG. 3 the absence of activity inthe TGA and DSC profiles below around 298° C. implies that the substanceshows good physical and chemical stability at normal operatingtemperatures.

[0087] As shown in the Examples, enthalpy of dissolution of Forms 1 and3 have been determined in certain organic solvents and accordingly anenthalpy of transition from Form 3 to Form 1 of 5.1-6.7 kJ/mol has beenestimated.

[0088] Thus we prefer compound of formula (I) in unsolvated Form 1 sincethis form appears to be thermodynamically most stable at ambienttemperature and also appears to be least susceptible to undesirablemoisture sorption (see results in Examples section).

[0089] Although use of a compound of formula (I) in solvated form is notpreferred, nevertheless we have surprisingly found that certain solvateforms have particularly attractive physicochemical properties whichmakes them useful as intermediates in the preparation of a compound offormula (I) in unsolvated form (eg by removal of solvent as a finalstep). For example we have discovered that certain stoichiometricsolvates can be isolated as solids in highly crystalline form. Thus wealso provide as an aspect of the invention:

[0090] Compound of formula (I) as the methylethylketone solvate

[0091] Compound of formula (I) as the isopropanol solvate

[0092] Compound of formula (I) as the tetrahydrofuran solvate

[0093] Compound of formula (I) as the acetone solvate.

[0094] In particular we provide the aforementioned solvates as solids incrystalline form. A further particular advantage of these solvates isthe fact that desolvation of the solvate (eg by heating) results information of the unsolvated form as the preferred Form 1. Theaforementioned solvates have relatively low toxicity and are suitablefor use in industrial scale manufacture. Compound of formula (I) as theDMF solvate which may also be isolated as a solid in crystalline form isalso of interest for use in onward processing to unsolvated Form 1.

[0095] The compound of formula (I) and solvates thereof may be preparedby the methodology described hereinafter, constituting a further aspectof this invention.

[0096] A process according to the invention for preparing a compound offormula (I) comprises alkylation of a thioacid of formula (II)

[0097] or a salt thereof.

[0098] In this process the compound of formula (II) may be reacted witha compound of formula FCH₂L wherein L represents a leaving group (eg ahalogen atom, a mesyl or tosyl group or the like) for example, anappropriate fluoromethyl halide under standard conditions. Preferably,the fluoromethyl halide reagent is bromofluoromethane.

[0099] As noted later, preferably the compound of formula (II) isemployed as a salt, particularly the salt with diisopropylethylamine.

[0100] In a preferred process for preparing the compound of formula (I),the compound of formula (II) or a salt thereof is treated withbromofluoromethane optionally in the presence of a phase transfercatalyst. A preferred solvent is methylacetate, or more preferablyethylacetate, optionally in the presence of water. The presence of waterimproves solubility of both starting material and product and the use ofa phase transfer catalyst results in an increased rate of reaction.Examples of phase transfer catalysts that may be employed include (butare not restricted to) tetrabutylammonium bromide, tetrabutylammoniumchloride, benzyltributylammonium bromide, benzyltributylammoniumchloride, benzyltriethylammonium bromide, methyltributylammoniumchloride and methyltrioctylammonium chloride. THF has also successfullybeen employed as solvent for the reaction wherein the presence of aphase transfer catalyst again provides a significantly faster reactionrate. Preferably the product present in an organic phase is washedfirstly with aqueous acid eg dilute HCl in order to remove aminecompounds such as triethylamine and diisopropylethylamine and then withaqueous base eg sodium bicarbonate in order to remove any unreactedprecursor compound of formula (II). As noted later, if the compound offormula (I) so produced in solution in ethylacetate is distilled andtoluene added, then unsolvated Form 1 crystallises out.

[0101] Compounds of formula (II) may be prepared from the corresponding17α-hydroxyl derivative of formula (III):

[0102] using for example, the methodology described by G. H. Phillippset al., (1994) Journal of Medicinal Chemistry, 37, 3717-3729. Forexample the step typically comprises the addition of a reagent suitablefor performing the esterification eg an activated derivative of 2-furoicacid such as an activated ester or preferably a 2-furoyl halide eg2-furoyl chloride (employed in at least 2 times molar quantity relativeto the compound of formula (III)) in the presence of an organic base egtriethylamine. The second mole of 2-furoyl chloride reacts with thethioacid moiety in the compound of formula (III) and needs to be removedeg by reaction with an amine such as diethylamine.

[0103] This method suffers disadvantages, however, in that the resultantcompound of formula (II) is not readily purified of contamination withthe by-product 2-furoyldiethylamide. We have therefore invented severalimproved processes for performing this conversion.

[0104] In a first such improved process we have discovered that by usinga more polar amine such as diethanolamine, a more water solubleby-product is obtained (in this case 2-furoyldiethanolamide) whichpermits compound of formula (II) or a salt thereof to be produced inhigh purity since the by-product can efficiently be removed by waterwashing.

[0105] Thus according to this aspect of the invention we provide aprocess for preparing a compound of formula (II) which comprises:

[0106] (a) reacting a compound of formula (III) with an activatedderivative of 2-furoic acid as in an amount of at least 2 moles of theactivated derivative per mole of compound of formula (III) to yield acompound of formula (IIA);

[0107] ;and

[0108] (b) removal of the sulphur-linked 2-furoyl moiety from compoundof formula (IIA) by reaction of the product of step (a) with an organicprimary or secondary amine base capable of forming a water soluble2-furoyl amide.

[0109] In two particularly convenient embodiments of this process wealso provide methods for the efficient purification of the end productwhich comprise either

[0110] (c1) when the product of step (b) is dissolved in a substantiallywater immiscible organic solvent, purifying the compound of formula (II)by washing out the amide by-product from step (b) with an aqueous wash,or

[0111] (c2) when the product of step (b) is dissolved in a watermiscible solvent, purifying the compound of formula (II) by treating theproduct of step (b) with an aqueous medium so as to precipitate out purecompound of formula (II) or a salt thereof.

[0112] In step (a) preferably the activated derivative of 2-furoic acidmay be an activated ester of 2-furoic acid, but is more preferably a2-furoyl halide, especially 2-furoyl chloride. A suitable solvent forthis reaction is ethylacetate or methylacetate (preferablymethylacetate) (when step (c1) may be followed) or acetone (when step(c2) may be followed). Normally an organic base eg triethylamine will bepresent. In step (b) preferably the organic base is diethanolamine. Thebase may suitably be dissolved in a solvent eg methanol. Generally steps(a) and (b) will be performed at reduced temperature eg between 0 and 5°C. In step (c1) the aqueous wash may be water, however the use of brineresults in higher yields and is therefore preferred. In step (c2) theaqueous medium is for example a dilute aqueous acid such as dilute HCl.

[0113] According to a related aspect of the invention we provide analternative process for preparing a compound of formula (II) whichcomprises:

[0114] (a) reacting a compound of formula (III) with an activatedderivative of 2-furoic acid in an amount of at least 2 moles ofactivated derivative per mole of compound of formula (III) to yield acompound of formula (IIA); and

[0115] (b) removal of the sulphur-linked 2-furoyl moiety from compoundof formula (IIA) by reaction of the product of step (a) with a furthermole of compound of formula (III) to give two moles of compound offormula (II).

[0116] In step (a) preferably the activated derivative of 2-furoic acidmay be an activated ester of 2-furoic acid, but is more preferably a2-furoyl halide, especially 2-furoyl chloride. A suitable solvent forhis step is acetone. Normally an organic base eg triethylamine will bepresent. In step (b) a suitable solvent is DMF or dimethylacetamide.Normally an organic base eg triethylamine will be present. Generallysteps (a) and (b) will be performed at reduced temperature eg between 0and 5° C. The product may be isolated by treatment with acid and washingwith water.

[0117] This aforementioned process is very efficient in that it does notproduce any furoylamide by-product (thus affording inter aliaenvironmental advantages) since the excess mole of furoyl moiety istaken up by reaction with a further mole of compound of formula (II) toform an additional mole of compound of formula (II).

[0118] Further general conditions for the conversion of compound offormula (III) to compound of formula (II) in the two processes justdescribed will be well known to persons skilled in the art.

[0119] According to a preferred set of conditions, however, we havefound that the compound of formula (II) may advantageously be isolatedin the form of a solid crystalline salt. The preferred salt is a saltformed with a base such as triethylamine, 2,4,6-trimethylpyridine,diisopropylethylamine or N-ethylpiperidine. Such salt forms of compoundof formula (II) are more stable, more readily filtered and dried and canbe isolated in higher purity than the free thioacid. The most preferredsalt is the salt formed with diisopropylethylamine. The triethylaminesalt is also of interest.

[0120] Compounds of formula (III) may be prepared in accordance withprocedures described in GB 2088877B.

[0121] Compounds of formula (III) may also be prepared by a processcomprising the following steps:

[0122] Step (a) comprises oxidation of a solution containing thecompound of formula (V). Preferably, step (a) will be performed in thepresence of a solvent comprising methanol, water, tetrahydrofuran,dioxan or diethylene glygol dimethylether. So as to enhance yield andthroughput, preferred solvents are methanol, water or tetrahydrofuran,and more preferably are water or tetrahydrofuran, especially water andtetrahydrofuran as solvent. Dioxan and diethylene glygol dimethyletherare also preferred solvents which may optionally (and preferably) beemployed together with water. Preferably, the solvent will be present inan amount of between 3 and 10 vol relative to the amount of the startingmaterial (1 wt.), more preferably between 4 and 6 vol., especially 5vol. Preferably the oxidising agent is present in an amount of 1-9 molarequivalents relative to the amount of the starting material. Forexample, when a 50% w/w aqueous solution of periodic acid is employed,the oxidising agent may be present in an amount of between 1.1 and 10wt. relative to the amount of the starting material (1 wt.), morepreferably between 1.1 and 3 wt., especially 1.3 wt. Preferably, theoxidation step will comprise the use of a chemical oxidising agent. Morepreferably, the oxidising agent will be periodic acid or iodic acid or asalt thereof. Most preferably, the oxidising agent will be periodic acidor sodium periodate, especially periodic acid. Alternatively (or inaddition), it will also be appreciated that the oxidation step maycomprise any suitable oxidation reaction, eg one which utilises airand/or oxygen. When the oxidation reaction utilises air and/or oxygen,the solvent used in said reaction will preferably be methanol.Preferably, step (a) will involve incubating the reagents at roomtemperature or a little warmer, say around 25° C. eg for 2 hours. Thecompound of formula (IV) may be isolated by recrystallisation from thereaction mixture by addition of an anti-solvent. A suitable anti-solventfor compound of formula (IV) is water. Surprisingly we have discoveredthat it is highly desirable to control the conditions under which thecompound of formula (IV) is precipitated by addition of anti-solvent egwater. When the recrystallisation is performed using chilled water (egwater/ice mixture at a temperature of 0-5° C.) although betteranti-solvent properties may be expected we have found that thecrystalline product produced is very voluminous, resembles a soft geland is very difficult to filter. Without being limited by theory webelieve that this low density product contains a large amount ofsolvated solvent within the crystal lattice. By contrast when conditionsof around 10° C. or higher are used (eg around ambient temperature) agranular product of a sand like consistency which is very easilyfiltered is produced. Under these conditions, crystallisation typicallycommences after around 1 hour and is typically completed within a fewhours (eg 2 hours). Without being limited by theory we believe that thisgranular product contains little or no solvated solvent within thecrystal lattice.

[0123] Step (b) will typically comprise the addition of a reagentsuitable for converting a carboxylic acid to a carbothioic acid eg usinghydrogen sulphide gas together with a suitable coupling agent egcarbonyldiimidazole (CDI) in the presence of a suitable solvent egdimethylformamide.

[0124] An alternative process for preparing a compound of formula (II)comprises treating a compound of formula (X) with a reagent suitable forconverting a carboxylic acid to a carbothioic acid eg using hydrogensulphide gas together with a suitable coupling agent such as CDI in thepresence of a suitable solvent eg DMF. Compounds of formula (X) may beprepared by methodology analogous to that described herein.

[0125] An alternative process for preparing a compound of formula (I) ora solvate thereof comprises reacting a compound of formula (VI)

[0126] with a fluorine source.

[0127] Examples of suitable sources of fluorine include fluoride (egsodium fluoride) or, more preferably, HF. The preferred reagent isaqueous HF. A solvent such as THF or DMF may be employed.

[0128] A compound of formula (VI) may be prepared by a processcomprising

[0129] (a) alkylating a compound of formula (VII)

[0130]  or a salt thereof;

[0131] (b) reacting a compound of formula (VIII)

[0132]  with an epoxide forming reagent; or

[0133] (c) esterifying a compound of formula (IX)

[0134] In process (a), analogous conditions to those described above forthe conversion of a compound of formula (II) to a compound of formula(I) may be employed. Typically compound of formula (VII) will be reactedwith a compound of formula FCH₂L wherein L represents a leaving group(eg a halogen atom, a mesyl or tosyl group or the like) for example, anappropriate fluoromethyl halide under standard conditions. Preferably,the fluoromethyl halide reagent is bromofluoromethane.

[0135] Process (b) is preferably performed in two steps: (i) formationof a halohydrin especially a bromohydrin (eg by reaction with bromodanor equivalent reagent), followed by (ii) treatment with base such assodium hydroxide so as to effect ring closure. The product of step (i)is a compound of formula (IXA) which is a novel intermediate that may beisolated, if desired:

[0136] wherein X represents halogen, especially Br.

[0137] In process (c), a suitable reagent would be an activatedderivative of 2-furoic acid such as an activated ester or preferably a2-furoyl halide eg 2-furoyl chloride in the presence of an organic baseeg triethylamine. This reaction may be performed at elevated temperatureeg around 60° C. or else at ambient temperature in the presence of anacylation catalyst eg dimethylamino pyridine (DMAP).

[0138] Compounds of formula (VII) may be prepared by a processcomprising esterification of a compound of formula (XI)

[0139] Analogous conditions to those described above for the conversionof a compound of formula (III) to a compound of formula (II) may beemployed. For example, a suitable reagent would be an activatedderivative of 2-furoic acid such as an activated ester or preferably a2-furoyl halide eg 2-furoyl chloride in the presence of an organic baseeg triethylamine. Compound of formula (XI) is known (J Labelled CompdRadiopharm (1997) 39(7) 567-584).

[0140] A compound of formula (VIII) may be prepared by a processcomprising

[0141] (a) alkylating a compound of formula (XII)

[0142]  or a salt thereof; or

[0143] (b) esterifying a compound of formula (XIII)

[0144] In process (a), analogous conditions to those described above forthe conversion of a compound of formula (II) to a compound of formula(I) may be employed. Typically compound of formula (XII) will be reactedwith a compound of formula FCH₂L wherein L represents a leaving group(eg a halogen atom, a mesyl or tosyl group or the like) for example, anappropriate fluoromethyl halide under standard conditions. Preferably,the fluoromethyl halide reagent is bromofluoromethane.

[0145] In process (b), analogous conditions to those employed above forthe conversion of a compound of formula (IX) to a compound of formula(VI) may be employed. For example, a suitable reagent would be anactivated derivative of 2-furoic acid such as an activated ester orpreferably a 2-furoyl halide eg 2-furoyl chloride in the presence of anorganic base eg triethylamine.

[0146] Compounds of formula (IX) and (XIII) may be prepared byalkylating the corresponding thioacids (XI) and (XIV) (defined below)using methodology analogous to that already described (eg by reactionwith a compound of formula FCH₂L wherein L represents a leaving group(eg a halogen atom, a mesyl or tosyl group or the like) for example, anappropriate fluoromethyl halide under standard conditions. Preferably,the fluoromethyl halide reagent is bromofluoromethane. The thioacid (XI)is a known compound (J Labelled Compd Radiopharm (1997) 39(7) 567-584).

[0147] Compound of formula (XII) may be prepared by a process comprisingesterifying a compound of formula (XIV):

[0148] or a salt thereof.

[0149] This process may be performed using methodology analogous to thatalready described. For example, a suitable reagent would be an activatedderivative of 2-furoic acid such as an activated ester or preferably a2-furoyl halide eg 2-furoyl chloride in the presence of an organic baseeg triethylamine.

[0150] Compounds of formula (XIV) may be prepared from the correspondingcarboxylic acid eg by a process analogous to that described above forthe conversion of a compound of formula (IV) to a compound of formula(III). The aforesaid corresponding carboxylic acid is known (Upjohn, WO90/15816).

[0151] A further alternative process for preparing a compound of formula(I) or a solvate thereof comprises deprotecting or unmasking a compoundof formula (I) in which the 11-β-hydroxy group is protected or masked. Afirst such process comprises deprotecting a compound of formula (XV)

[0152] wherein P represents a hydroxy protecting group.

[0153] Examples of hydroxy protecting groups P are described inProtective Groups in Organic Chemistry Ed J F W McOmie (Plenum Press1973) or Protective Groups in Organic Synthesis by Theodora W Green(John Wiley and Sons, 1991).

[0154] Examples of suitable hydroxy protecting groups P include groupsselected from carbonate, alkyl (eg t-butyl or methoxymethyl), aralkyl(eg benzyl, p-nitrobenzyl, diphenylmethyl or triphenylmethyl),heterocyclic groups such as tetrahydropyranyl, acyl (eg acetyl orbenzyl) and silyl groups such as trialkylsilyl (egt-butyldimethylsilyl). The hydroxy protecting groups may be removed byconventional techniques. Thus, for example, carbonate may be removed bytreatment with base and alkyl, silyl, acyl and heterocyclic groups maybe removed by solvolysis eg by hydrolysis under acid or basicconditions. Aralkyl groups such as triphenylmethyl may similarly beremoved by solvolysis eg by hydrolysis under acidic conditions. Aralkylgroups such as benzyl or p-nitrobenzyl may be cleaved by hydrogenolysisin the presence of a Noble metal catalyst such as palladium on charcoal.p-Nitrobenzyl may also be cleaved by photolysis.

[0155] The 11-β-hydroxy group may be masked as a carbonyl group. Thus asecond such process comprises reduction of a compound of formula (XVI)

[0156] Reduction to the compound of formula (I) may be achieved eg bytreatment with a hydride reducing agent such as borohydride eg sodiumborohydride.

[0157] The 11-ketone (XVI) may also be masked. Examples of maskedderivatives of compound of formula (XVI) include (i) ketal derivativeseg ketals formed by treatment of the compound of formula (XVI) with analcohol eg methanol, ethanol or ethan-1,2-diol, (ii) dithioketalderivatives eg dithioketals formed by treatment of the compound offormula (XVI) with a thiol eg methanethiol, ethanethiol orethan-1,2-dithiol, (iii) monothioketal derivatives eg monothioketalsformed by treatment of the compound of formula (XVI) with eg1-hydroxy-ethane-2-thiol, (iv) derivatives formed by treatment of thecompound of formula (XVI) with an alcoholamine eg ephedrine, (v) iminesformed by treatment of the compound of formula (XVI) with amines, (vi)oximes formed by treatment of compounds of formula (XVI) withhydroxylamines. We claims such derivatives of compound of formula (XVI)as an aspect of the invention.

[0158] These masked derivatives may be converted back to the ketone byconventional means eg ketals, imines and oximes are converted tocarbonyl by treatment with dilute acid and dithioketals are converted tothe ketone by a variety of methods as described by P. C. Bulman Page etal (1989), Tetrahedron, 45, 7643-7677 and references therein.

[0159] Compounds of formula (XV) may be prepared by a process comprising

[0160] (a) alkylating a compound of formula (XVII)

[0161]  or a salt thereof wherein P represents a hydroxy protectinggroup; or

[0162] (b) esterifying a compound of formula (XVIII)

[0163] In step (a), analogous conditions to those described above forthe conversion of a compound of formula (II) to a compound of formula(I) may be employed. Typically compound of formula (XVII) will bereacted with a compound of formula FCH₂L wherein L represents a leavinggroup (eg a halogen atom, a mesyl or tosyl group or the like) forexample, an appropriate fluoromethyl halide under standard conditions.Preferably, the fluoromethyl halide reagent is bromofluoromethane.

[0164] In step (b), analogous conditions to those employed above for theconversion of a compound of formula (IX) to a compound of formula (VI)may be employed. For example, a suitable reagent would be an activatedderivative of 2-furoic acid such as an activated ester or preferably a2-furoyl halide eg 2-furoyl chloride in the presence of an organic baseeg triethylamine.

[0165] Compound of formula (XVIII) may be prepared by alkylating thecorresponding thioacid using methodology analogous to that alreadydescribed (eg by reaction with a compound of formula FCH₂L wherein Lrepresents a leaving group (eg a halogen atom, a mesyl or tosyl group orthe like) for example, an appropriate fluoromethyl halide under standardconditions. Preferably, the fluoromethyl halide reagent isbromofluoromethane. The corresponding thioacids are known compounds ormay be prepared by standard methodology. Compound of formula (XVIII) mayalternatively be prepared by protection of the corresponding hydroxyderivative.

[0166] Compound of formula (XVII) may be prepared by a processcomprising esterifying a compound of formula (XIX)

[0167] or a salt thereof wherein P represents a hydroxy protectinggroup.

[0168] This process may be performed using methodology analogous to thatalready described for the conversion of compounds of formula (III) to(II). For example, a suitable reagent would be an activated derivativeof 2-furoic acid such as an activated ester or preferably a 2-furoylhalide eg 2-furoyl chloride in the presence of an organic base egtriethylamine.

[0169] Compounds of formula (XIX) may be prepared by protecting thecorresponding hydroxy derivative (III), having first protected thethioacid which would then be deprotected.

[0170] Compounds of formula (XVI) may be prepared by a processcomprising

[0171] (a) alkylating a compound of formula (XX)

[0172]  or a salt thereof or a derivative wherein the 11-carbonyl groupis masked; or

[0173] (b) esterifying a compound of formula (XXI)

[0174]  or a derivative wherein the 11-carbonyl group is masked.

[0175] In step (a), analogous conditions to those described above forthe conversion of a compound of formula (III) to a compound of formula(II) may be employed. Typically compound of formula (XX) will be reactedwith a compound of formula FCH₂L wherein L represents a leaving group(eg a halogen atom, a mesyl or tosyl group or the like) for example, anappropriate fluoromethyl halide under standard conditions. Preferably,the fluoromethyl halide reagent is bromofluoromethane.

[0176] In step (b), analogous conditions to those employed above for theconversion of a compound of formula (IX) to a compound of formula (VI)may be employed. For example, a suitable reagent would be an activatedderivative of 2-furoic acid such as an activated ester or preferably a2-furoyl halide eg 2-furoyl chloride in the presence of an organic baseeg triethylamine.

[0177] Compound of formula (XXI) or a derivative thereof wherein the11-ketone group is masked may be prepared by alkylating thecorresponding thioacid using methodology analogous to that alreadydescribed (eg by reaction with a compound of formula FCH₂L wherein Lrepresents a leaving group (eg a halogen atom, a mesyl or tosyl group orthe like) for example, an appropriate fluoromethyl halide under standardconditions. Preferably, the fluoromethyl halide reagent isbromofluoromethane. The corresponding thioacids are known compounds ormay be prepared from the corresponding carboxylic acids by methodsanalogous to those previously described.

[0178] Compound of formula (XX) may be prepared by a process comprisingesterifying a compound of formula (XXII)

[0179] or a derivative thereof wherein the 11-ketone group is masked.

[0180] This process may be performed using methodology analogous to thatalready described. For example, a suitable reagent would be an activatedderivative of 2-furoic acid such as an activated ester or preferably a2-furoyl halide eg 2-furoyl chloride in the presence of an organic baseeg triethylamine.

[0181] Compounds of formula (XXII) and derivatives thereof wherein the11-ketone is masked may be prepared by oxidation of the correspondinghydroxy derivative (IV) followed by masking of the ketone and subsequentconversion of the carboxylic acid group to the thioacid (see egconversion of compounds of formula (IV) to (III).

[0182] A further alternative process for the preparation of compounds offormula (I) or a solvate thereof comprises reaction of a compound offormula (XXIII)

[0183] wherein L represents a leaving group (eg halide other thanfluoride such as chloride, iodide or a sulphonate ester such mesylate,tosylate, triflate) with a fluorine source.

[0184] Preferably the fluorine source is fluoride ion eg KF. Furtherdetails for this conversion may be obtained by reference to G. H.Phillipps et al., (1994) Journal of Medicinal Chemistry, 37, 3717-3729or J Labelled Compd Radiopharm (1997) 39(7) 567-584).

[0185] Compounds of formula (XXIII) may be prepared by methods analogousto those described herein. Corresponding novel intermediates of formula(VI), (VIII), (IX), (IXA), (XV) and (XVI) wherein the —CH2F moiety isreplaced with a —CH2L moiety (wherein L represents a leaving group otherthan fluorine) are claimed as an aspect of the invention.

[0186] A further alternative process for the preparation of compounds offormula (I) or a solvate thereof comprises deprotection or unmasking ofa derivative of a compound of formula (I) in which the 3-carbonyl groupis protected or masked.

[0187] The 3-carbonyl group may be masked in a manner analogous to thatdescribed above in relation to masking of the 11-carbonyl position. Thusthe 3-carbonyl may be masked eg as a ketal, monothioketal, dithioketal,derivative with an alcoholamine, oxime or imine. The carbonyl group maybe recovered by conventional means eg ketals are converted to carbonylby treatment with dilute acid and dithioketals are converted to theketone by a variety of methods as described by P. C. Bulman Page et al(1989), Tetrahedron, 45, 7643-7677 and references therein.

[0188] Certain intermediate compounds are new and we provide these,together where appropriate with their salts and solvates, as an aspectof the invention.

[0189] As noted above, we provide as a particular aspect of theinvention a process for preparing a compound of formula (I) inunsolvated form which comprises:

[0190] (a) Crystallising the compound of formula (I) in the presence ofa non-solvating solvent such as ethanol, methanol, water, ethyl acetate,toluene, methylisobutylketone or mixtures thereof; or

[0191] (b) Desolvating a compound of formula (I) in solvated form (eg inthe form of a solvate with acetone, isopropanol, methylethylketone, DMFor tetrahydrofuran) eg by heating.

[0192] In step (b) the desolvation will generally be performed at atemperature exceeding 50° C. preferably at a temperature exceeding 100°C. Generally heating will be performed under vacuum.

[0193] There is also provided a compound of formula (I) in unsolvatedform obtainable by the aforementioned process.

[0194] There is also provided as a particular aspect of the invention aprocess for preparing a compound of formula (I) as unsolvated Form 1polymorph which comprises dissolving compound of formula (I) inmethylisobutylketone, ethyl acetate or methyl acetate and producingcompound of formula (I) as unsolvated Form 1 by addition of anon-solvating anti-solvent such as iso-octane or toluene.

[0195] According to a first preferred embodiment of this process thecompound of formula (I) may be dissolved in ethyl acetate and compoundof formula (I) as unsolvated Form 1 polymorph may be obtained byaddition of toluene as anti-solvent. In order to improve the yield,preferably the ethyl acetate solution is hot and once the toluene hasbeen added the mixture is distilled to reduce the content of ethylacetate.

[0196] According to a second preferred embodiment of this process thecompound of formula (I) may be dissolved in methylisobutylketone andcompound of formula (I) as unsolvated Form 1 polymorph may be obtainedby addition of isooctane as anti-solvent.

[0197] There is also provided a compound of formula (I) as unsolvatedForm 1 polymorph obtainable by the aforementioned processes.

[0198] A process for preparing a compound of formula (I) as unsolvatedForm 2 polymorph comprises dissolving compound of formula (I) inunsolvated form in methanol or dry dichloromethane and recrystallisingthe compound of formula (I) as unsolvated Form 2 polymorph. Typicallythe compound of formula (I) will be dissolved in hot in methanol or drydichloromethane and allowed to cool.

[0199] There is also provided a compound of formula (I) as unsolvatedForm 2 polymorph obtainable by the aforementioned process.

[0200] A process for preparing a preparing a compound of formula (I) asunsolvated Form 3 polymorph comprises dissolving compound of formula (I)in particular as the acetone solvate in dichloromethane in the presenceof water (typically 1-3% water by volume) and recrystallising thecompound of formula (I) as unsolvated Form 3 polymorph.

[0201] There is also provided a compound of formula (I) as unsolvatedForm 3 polymorph obtainable by the aforementioned process.

[0202] The advantages of the compound of formula (I) and/or its solvatesor polymorphs may include the fact that the substance appears todemonstrate excellent anti-inflammatory properties, with predictablepharmacokinetic and pharmacodynamic behaviour, with an attractiveside-effect profile, long duration of action, and is compatible with aconvenient regime of treatment in human patients, in particular beingamendable to once-per day dosing. The advantages may be appreciated inparticular when the compound of formula (I) and/or its solvates orpolymorphs are employed in combination with a the long-actingβ₂-adrenoreceptor agonist. Further advantages may include the fact thatthe substance has desirable physical and chemical properties which allowfor ready manufacture and storage.

BRIEF DESCRIPTION OF THE FIGURES

[0203]FIG. 1: Overlay of the XRPD profiles of Form 1, Form 2 and Form 3polymorphs of unsolvated Compound of formula (I).

[0204]FIG. 2: Overlay of the XRPD profiles of Form 1, Form 2 and a 50:50mixture of Form 1 and Form 2 polymorphs of unsolvated Compound offormula (I) together with the time dependence of the profile of the50:50 mixture of Form 1 and Form 2.

[0205]FIG. 3: DSC and TGA profiles of Form 1 polymorph of UnsolvatedCompound of formula (I).

[0206]FIG. 4: Temperature dependence of the XRPD profile of Compound offormula (I) Unsolvated Form 3 obtained at 5 timepoints.

[0207]FIG. 5: Temperature and time profile for the XRPD experiments ofFIG. 4.

[0208] The following non-limiting Examples illustrate the invention:

EXAMPLES General

[0209]¹H-nmr spectra were recorded at 400 MHz and the chemical shiftsare expressed in ppm relative to tetramethylsilane. The followingabbreviations are used to describe the multiplicities of the signals: s(singlet), d (doublet), t (triplet), q (quartet), m (multiplet), dd(doublet of doublets), ddd (doublet of doublet of doublets), dt (doubletof triplets) and b (broad). Biotage refers to prepacked silica gelcartridges containing KP-Sil run on flash 12i chromatography module.LCMS was conducted on a Supelcosil LCABZ+PLUS column (3.3 cm×4.6 mm ID)eluting with 0.1% HCO₂H and 0.01 M ammonium acetate in water (solventA), and 0.05% HCO₂H 5% water in acetonitrile (solvent B), using thefollowing elution gradient 0-0.7 min 0% B, 0.7-4.2 min 100% B, 4.2-5.3min 0% B, 5.3-5.5 min 0% B at a flow rate of 3 ml/min. The mass spectrawere recorded on a Fisons VG Platform spectrometer using electrospraypositive and negative mode (ES+ve and ES−ve).

[0210] DSC and TGA profiles were obtained using a Netzsch STA449Csimultaneous thermal analyser using an unsealed pan with nitrogen gasflow and a thermal gradient of 10° C./min.

[0211] The moisture sorption characteristics were obtained using a HidenIgasorb water sorption microbalance. The programme provides for stepwiseincrease in relative humidity (RH) from 0 to 90% RH and then decreaseback to 0% RH in steps of 10% RH.

[0212] The XRPD analysis shown in FIG. 1 and 2 were performed on aPhillips X'pert MPD powder diffractometer, serial number DY667. Themethod runs from 2 to 45 degrees 2Theta with 0.02 degree 2Theta stepsize and a 1 second collection time at each step. The XRPD analysisshown in FIG. 4 employed the same instrument with an Anton Parr TTKthermal accessory using a method running from 2 to 35 degrees 2Thetawith 0.04 degree 2Theta step size and a 1 second collection time.

Intermediates Intermediate 1: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid

[0213] A solution of 6α, 9α-difluoro-11β,17α-dihydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid(prepared in accordance with the procedure described in GB 2088877B) (18g, 43.64 mmol) in anhydrous dichloromethane (200 ml) and triethylamine(15.94 ml, 114 mmol) was treated at <5° C. with a solution of 2-furoylchloride (11.24 ml, 114 mmol) in anhydrous dichloromethane (100 ml) overapproximately 40 min. The solution was stirred at <5° C. for 30 min. Theresulting solid was collected by filtration, washed successively with3.5% aqueous sodium hydrogen carbonate solution, water, 1M hydrochloricacid, and water and dried in vacuo at 60° C. to give a cream colouredsolid. The dichloromethane filtrate was washed successively with 3.5%sodium hydrogen carbonate solution, water, 1M hydrochloric acid, water,dried (Na₂SO₄) and evaporated to give a cream coloured solid which wascombined with that isolated above. The combined solids (26.9 g) weresuspended in acetone (450 ml) and stirred. Diethylamine (16.8 ml, 162mmol) was added and the mixture stirred at room temperature for 4.5 h.The mixture was concentrated and the precipitate collected by filtrationand washed with a little acetone. The washings and filtrate werecombined, concentrated and loaded onto a silica gel Biotage column whichwas eluted with 24:1 chloroform:methanol. Fractions which contained themore polar component were combined and evaporated to give a creamcoloured solid. This was combined with the solid isolated above anddried in vacuo to give a pale beige coloured solid (19.7 g). This wasdissolved in warm water, the pH adjusted to 2 with concentratedhydrochloric acid and the mixture extracted with ethyl acetate. Theorganic extract was dried (Na₂SO₄) and evaporated to give, after dryingat 50° C., the title compound as a cream coloured solid (18.081 g, 82%):LCMS retention time 3.88 min, m/z 507 MH⁺, NMR δ (CDCl₃) includes 7.61(1H, m), 7.18-7.12 (2H, m), 6.52 (1H, dd, J 4, 2 Hz), 6.46 (1H, s), 6.41(1H, dd, J 10, 2 Hz), 5.47 and 5.35 (1H, 2m), 4.47 (1H, bd, J 9 Hz),3.37 (1H, m), 1.55 (3H, s), 1.21 (3H, s), 1.06 (3H, d, J 7 Hz).

Intermediate 1: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid (first alternative method)

[0214] A stirred suspension of 6α, 9α-difluoro-11β,17α-dihydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid(prepared in accordance with the procedure described in GB 2088877B) (1wt, 49.5 g) in acetone (10 vol) is cooled to 0-5° C. and treated withtriethylamine (0.51 wt, 2.1 eq), keeping the temperature below 5° C.,and stirred for 5 min at 0-5° C. 2-Furoyl chloride (0.65 wt, 2.05 eq) isthen added over a minimum of 20 min, maintaining a reaction temperatureat 0-5° C. The reaction is stirred for 30 min at 0-5° C. then sampledfor analysis by HPLC. A solution of diethanolamine (1.02 wt, 4 eq) inmethanol (0.8 vol) is added over ca 15 min followed by a line wash ofmethanol (0.2 vol) and the reaction stirred at 0-5° C. for 1 h. Thereaction is again sampled for analysis by HPLC then warmed toapproximately 20° C. and treated with water (1.1 wt). The reactionmixture is then treated with a solution of HCl (SG1.18 (11.5M), 1 vol)in water (10 vol) over ca 20 min maintaining a reaction temperaturebelow 25° C. The suspension is stirred at 20-23° C. for at least 30minutes then filtered. The filter cake is washed with water (3×2 vol).The product is dried in vacuo at approximately 60° C. overnight to givethe title compound as a white solid (58.7 g, 96.5%).

Intermediate 1: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid (second alternative method)

[0215] A stirred suspension of 6α, 9α-difluoro-11β,17α-dihydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid(prepared in accordance with the procedure described in GB 2088877B) (1wt, 49.5 g) in acetone (10 vol) is cooled to 0-5° C. and treated withtriethylamine (0.51 wt, 2.1 eq), keeping the temperature below 5° C.,and stirred for 5 min at 0-5° C. 2-Furoyl chloride (0.65 wt, 2.05 eq) isthen added over a minimum of 20 min, maintaining a reaction temperatureat 0-5° C. The reaction mixture is stirred for at least 30 minutes anddiluted with water (10 vol) maintaining a reaction temperature in therange 0-5° C. The resultant precipitate is collected by filtration andwashed sequentially with acetone/water (50/50 2 vol) and water (2×2vol). The product is dried under vacuum at approximately 55° C.overnight to leave6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-ylS-(2-furanylcarbonyl) thioanhydride as a white solid (70.8 g, 98.2%)(NMR δ (CD₃CN) 0.99 (3H, d) (J=7.3 Hz), 1.24 (3H, s), 1.38 (1H, m)(J=3.9 Hz), 1.54 (3H, s), 1.67 (1H, m), 1.89 (1H, broad d) (J=15.2 Hz),1.9-2.0 (1H, m), 2.29-2.45 (3H, m), 3.39 (1H, m), 4.33 (1H, m), 4.93(1H, broad s), 5.53 (1H, ddd) (J=6.9,1.9 Hz; J_(HF)=50.9 Hz), 6.24 (1H,m), 6.29 (1H, dd) (J=10.3, 2.0 Hz), 6.63 (2H, m), 7.24-7.31 (3H, m),7.79 (1H, dd) (J=<1 Hz), 7.86 (1H, dd) (J=<1 Hz)). A portion of theproduct (0.56 g) is mixed with 6α, 9α-difluoro-11β,17α-dihydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid(0.41 g) in a 1:1 molar ratio in DMF (10 volumes wrt total steroidinput). The reaction mixture is treated with triethylamine(approximately 2.1 equivalents) and the mixture is stirred atapproximately 20° C. for approximately 6 hours. Water (50 vol)containing excess conc HCl (0.5 vol) is added to the reaction mixtureand the resultant precipitate collected by filtration. The bed is washedwith water (2×5 vol) and dried in vacuo at approximately 55° C.overnight to leave the title compound as a white solid (0.99 g, 102%).

Intermediate 1A: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid diisopropylethylamine salt

[0216] A stirred suspension of 6α, 9α-difluoro-11β,17α-dihydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid(prepared in accordance with the procedure described in GB 2088877B)(49.5 g) in methylacetate (500 ml) is treated with triethylamine (35 ml)maintaining a reaction temperature in the range 0-5° C. 2-Furoylchloride (25 ml) is added and the mixture stirred at 0-5° C. for 1 hour.A solution of diethanolamine (52.8 g) in methanol (50 ml) is added andthe mixture stirred at 0-5° C. for at least 2 hours. Dilute hydrochloricacid (approx 1M, 550 ml) is added maintaining a reaction temperaturebelow 15° C. and the mixture stirred at 15° C. The organic phase isseparated and the aqueous phase is back extracted with methyl acetate(2×250 ml). All of the organic phases are combined, washed sequentiallywith brine (5×250 ml) and treated with di-isopropylethylamine (30 ml).The reaction mixture is concentrated by distillation at atmosphericpressure to an approximate volume of 250 ml and cooled to 25-30° C.(crystallisation of the desired product normally occurs duringdistillation/subsequent cooling). Tertiary butyl methyl ether (TBME)(500 ml) is added, the slurry further cooled and aged at 0-5° C. for atleast 10 minutes. The product is filtered off, washed with chilled TBME(2×200 ml) and dried under vacuum at approximately 40-50° C. (75.3 g,98.7%). NMR (CDCl₃) δ: 7.54-7.46 (1H, m), 7.20-7.12 (1H, dd), 7.07-6.99(1H, dd), 6.48-6.41 (2H, m), 6.41-6.32 (1H, dd), 5.51-5.28 (1H, dddd²J_(H-F) 50 Hz), 4.45-4.33(1H, bd), 3.92-3.73 (3H, bm), 3.27-3.14 (2H,q), 2.64-2.12 (5H, m), 1.88-1.71 (2H, m), 1.58-1.15 (3H, s), 1.50-1.38(15H, m), 1.32-1.23 (1H, m), 1.23-1.15 (3H s), 1.09-0.99 (3H, d)

Intermediate 1B: 6α,9α-Difluoro-17α[-(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid triethylamine salt

[0217] A stirred suspension of Intermediate 1 (30 g) in ethylacetate(900 ml) is treated with triethylamine (1.05 molar equivalents, 8.6 ml)and the mixture is stirred at approximately 20° C. for 1.5 hours. Theprecipitate is filtered off, washed with ethylacetate (2×2 vol) anddried in vacuo at 45° C. for 18 hours to give title compound as a whitesolid (28.8 g, 80%). NMR (CDCl₃) δ: 7.59-7.47 (1H, m), 7.23-7.13 (1H,dd), 7.08-6.99 (1H, d), 6.54-6.42 (2H, m), 6.42-6.32 (1H, dd), 5.55-5.26(1H, dddd²J_(H-F) 50 Hz), 4.47-4.33(1H, bd), 3.88-3.70 (1H, bm),3.31-3.09 (6H, q), 2.66-2.14 (5H, m), 1.93-1.69 (2H, m), 1.61-1.48 (3H,s), 1.43-1.33 (9H, t), 1.33-1.26 (1H, m), 1.26-1.15 (3H s), 1.11-0.97(3H, d).

EXAMPLES Example 1 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester Unsolvated Form 1

[0218] A suspension of Intermediate 1 (2.5 g, 4.94 mmol) was dissolvedin anhydrous N, N-dimethylformamide (25 ml) and sodium hydrogencarbonate (465 mg, 5.53 mmol) was added. The mixture was stirred at −20°C. and bromofluoromethane (0.77 ml, 6.37 mmol) was added and the mixturewas stirred at −20° C. for 2 h. Diethylamine (2.57 ml, 24.7 mmole) wasadded and the mixture stirred at −20° C. for 30 min. The mixture wasadded to 2M hydrochloric acid (93 ml) and stirred for 30 min. Water (300ml) was added and the precipitate was collected by filtration, washedwith water and dried in vacuo at 50° C. to give a white solid which wasrecrystallised from acetone/water (to yield the acetone solvate of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17α-carbothioicacid S-fluoromethyl ester) and dried in vacuo at 50° C. to give thetitle compound (2.351 g, 88%): LCMS retention time 3.66 min, m/z 539MH⁺, NMR δ (CDCl₃) includes 7.60 (1H, m), 7.18-7.11 (2H, m), 6.52 (1H,dd, J 4.2 Hz), 6.46 (1H, s), 6.41 (1H, dd, J 10, 2 Hz), 5.95 and 5.82(2H dd, J 51, 9 Hz), 5.48 and 5.35 (1H, 2m), 4.48 (1H, m), 3.48 (1H, m),1.55 (3H, s), 1.16 (3H, s), 1.06 (3H, d, J 7 Hz).

[0219] Pharmacological Activity

[0220] In vitro Pharmacological Activity

[0221] Pharmacological activity was assessed in a functional in vitroassay of glucocorticoid agonist activity which is generally predictiveof anti-inflammatory or anti-allergic activity in vivo.

[0222] For the experiments in this section, compound of formula (I) wasused as unsolvated Form 1.

[0223] The functional assay was based on that described by K. P. Ray etal., Biochem J. (1997), 328, 707-715. A549 cells stably transfected witha reporter gene containing the NF-κB responsive elements from the ELAMgene promoter coupled to sPAP (secreted alkaline phosphatase) weretreated with test compounds at appropriate doses for 1 hour at 37° C.The cells were then stimulated with tumour necrosis factor (TNF, 10ng/ml) for 16 hours, at which time the amount of alkaline phosphataseproduced is measured by a standard colourimetric assay. Dose responsecurves were constructed from which EC₅₀ values were estimated.

[0224] In this test the compound of Example 1 showed an EC₅₀ value of <1nM.

[0225] The glucocorticoid receptor (GR) can function in at least twodistinct mechanisms, by upregulating gene expression through the directbinding of GR to specific sequences in gene promotors, and bydownregulating gene expression that is being driven by othertranscription factors (such as NFκB or AP-1) through their directinteraction with GR.

[0226] In a variant of the above method, to monitor these functions, tworeporter plasmids have been generated and introduced separately intoA549 human lung epithelial cells by transfection. The first cell linecontains the firefly luciferase reporter gene under the control of asynthetic promoter that specifically responds to activation of thetranscription factor NFκB when stimulated with TNFα. The second cellline contains the renilla luciferase reporter gene under the control ofa synthetic promotor that comprises 3 copies of the consensusglucocorticoid response element, and which responds to directstimulation by glucocorticoids. Simultaneous measurement oftransactivation and transrepression was conducted by mixing the two celllines in a 1:1 ratio in 96 well plate (40,000 cells per well) andgrowing overnight at 37° C. Test compounds were dissolved in DMSO, andadded to the cells at a final DMSO concentration of 0.7%. Afterincubation for 1 h 0.5 ng/ml TNFα (R&D Systems) was added and after afurther 15 hours at 37° C., the levels of firefly and renilla luciferasewere measured using the Packard Firelite kit following themanufacturers' directions. Dose response curves were constructed fromwhich EC₅₀ values were determined. Transactivation (GR) Transrepression(NFκB) ED₅₀ (nM) ED₅₀ (nM) Compound of Formula 0.06 0.20 (I) Metabolite(X) >250 >1000 Fluticasone propionate 0.07 0.16

[0227] In vivo Pharmacological Activity

[0228] Pharmacological activity in vivo was assessed in an ovalbuminsensitised Brown Norway rat eosinophilia model. This model is designedto mimic allergen induced lung eosinophilia, a major component of lunginflammation in asthma.

[0229] For the experiments in this section, compound of formula (I) wasused as unsolvated Form 1.

[0230] Compound (I) produced dose dependant inhibition of lungeosinophilia in this model after dosing as an intra-tracheal (IT)suspension in saline 30 min prior to ovalbumin challenge. Significantinhibition is achieved after a single dose of 30 μg of compound (I) andthe response was significantly (p=0.016) greater than that seen with anequivalent dose of fluticasone propionate in the same study (69%inhibition with compound (I) vs 41% inhibition with fluticasonepropionate).

[0231] In a rat model of thymus involution 3 daily IT doses of 100 μg ofcompound (I) induced significantly smaller reductions in thymus weight(p=0.004) than an equivalent dose of fluticasone propionate in the samestudy (67% reduction of thymus weight with compound (I) vs 78% reductionwith fluticasone propionate).

[0232] Taken together these results indicate a superior therapeuticindex for compound (I) compared to fluticasone propionate.

[0233] In vitro Metabolism in Rat and Human Hepatocytes

[0234] Incubation of compound (I) with rat or human hepatocytes showsthe compound to be metabolised in an identical manner to fluticasonepropionate with the 17-β carboxylic acid (X) being the only significantmetabolite produced. Investigation of the rate of appearance of thismetabolite on incubation of compound (I) with human hepatocytes (37° C.,10 μM drug concentration, hepatocytes from 3 subjects, 0.2 and 0.7million cells/mL) shows compound (I) to be metabolised ca. 5-fold morerapidly than fluticasone propionate:— 17-β acid metabolite productionSubject Cell density (pmol/h) number (million cells/mL) Compound (I)Fluticasone propionate 1 0.2 48.9 18.8 1 0.7 73.3 35.4 2 0.2 118 9.7 20.7 903 23.7 3 0.2 102 6.6 3 0.7 580 23.9

[0235] Median metabolite production 102-118 pmol/h for compound (I) and18.8-23.0 pmol/h for fluticasone propionate.

[0236] Pharmacokinetics After Intravenous (IV) and Oral Dosing in Rats

[0237] Compound (I) was dosed orally (0.1 mg/kg) and IV (0.1 mg/kg) tomale Wistar Han rats and pharmacokinetic parameters determined. Compound(I) showed negligible oral bioavailability (0.9%) and plasma clearanceof 47.3 mL/min/kg, approaching liver blood flow (plasma clearance offluticasone propionate=45.2 mL/min/kg).

[0238] Pharmacokinetics After Intra-tracheal Dry Powder Dosing in thePig.

[0239] Anaesthetised pigs (2) were dosed intra-tracheally with ahomogenous mixture of compound (I) (1 mg) and fluticasone propionate (1mg) as a dry powder blend in lactose (10% w/w). Serial blood sampleswere taken for up to 8 h following dosing. Plasma levels of compound (I)and fluticasone propionate were determined following extraction andanalysis using LC-MS/MS methodology, the lower limits of quantitation ofthe methods were 10 and 20 pg/mL for compound (I) and fluticasonepropionate respectively. Using these methods compound (I) wasquantifiable up to 2 hours after dosing and fluticasone propionate wasquantifiable up to 8 hours after dosing. Maximum plasma concentrationswere observed for both compounds within 15 min after dosing. Plasmahalf-life data obtained from IV dosing (0.1 mg/kg) was used to calculateAUC (0-inf) values for compound (I). This compensates for the plasmaprofile of Compound (I) only being defined up to 2 hours after an ITdose and removes any bias due to limited data between compound (I) andfluticasone propionate.

[0240] C_(max) and AUC (0-inf) values show markedly reduced systemicexposure to compound (I) compared to fluticasone propionate:— Cmax(pg/mL) AUC (0-inf) (hr. pg/mL) Pig 1 Pig 2 Pig 1 Pig 2 Compound ofFormula (I) 117  81 254 221 Fluticasone propionate 277 218 455 495

[0241] The pharmacokinetic parameters for both compound (I) andfluticasone propionate were the same in the anaesthetised pig followingintravenous administration of a mixture of the two compounds at 0.1mg/kg. The clearance of these two glucocorticoids is similar is thisexperimental pig model.

Example 1 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester Unsolvated Form 1 (First Alternative Method)

[0242] A mobile suspension of Intermediate 1A (12.61 g, 19.8 mmol;equivalent to 10 g of Intermediate 1) in ethyl acetate (230 ml) andwater (50 ml) is treated with a phase transfer catalyst(benzyltributylammonium chloride, 10 mol %), cooled to 3° C. and treatedwith bromofluoromethane (1.10 ml, 19.5 mmol, 0.98 equivalents), washingin with prechilled (0° C.) ethyl acetate (EtOAc) (20 ml). The suspensionis stirred overnight, allowing to warm to 17° C. The aqueous layer isseparated and the organic phase is sequentially washed with 1M HCl (50ml), 1% w/v NaHCO₃ solution (3×50 ml) and water (2×50 ml). Theethylacetate solution is distilled at atmospheric pressure until thedistillate reaches a temperature of approximately 73° C. at which pointtoluene (150 ml) is added. Distillation is continued at atmosphericpressure until all remaining EtOAc has been removed (approximatedistillate temperature 103° C.). The resultant suspension is cooled andaged at <10° C. and filtered off. The bed is washed with toluene (2×30ml) and the product oven dried under vacuum at 60° C. to constant weightto yield the title compound (8.77 g, 82%).

Example 1 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0243] Unsolvated Form 1 (Second Alternative Method)

[0244] A suspension of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester acetone solvate (prepared eg according toExample 11) (50.0 g) in acetone (1500 ml) and water (75 ml) was heatedto reflux. The resultant mixture was clarified by hot filtration(Whatman 54 filter paper) during which time some solid crystallised inthe filtrate. Further acetone (200 ml) was added to the filtrate givinga bright solution at reflux. The solution was distilled at atmosphericpressure until turbidity was noted whilst at reflux (approx 750 mlsolvent collected). Toluene (1000 ml) was added to the hot solution anddistillation at atmospheric pressure was continued givingcrystallisation at a temperature of approximately 98° C. Distillation ofsolvent was continued until a reaction temperature of 105° C. wasachieved (approximately 945 ml solvent collected). The mixture wascooled to ambient temperature, further cooled and aged at <10° C. for 10minutes. The product was filtered off, washed with toluene (150 ml) andsucked dry. The product was dried at approximately 60° C. under vacuumfor 16 h to leave the title compound as a dense white solid (37.8 g,83.7%).

[0245] The XRPD pattern of Example 1 product is shown in FIG. 1. The DSCand TGA profiles are shown in FIG. 3.

Example 2 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0246] Unsolvated Form 2

[0247] A suspension of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (prepared for example according to Example 1,first method) (6.0 g) in dichloromethane (180 ml) was heated to refluxgiving a bright solution. The solution was clarified by hot filtration(Whatman 54 filter paper) and the solution was distilled at atmosphericpressure (approx 100 ml solvent collected) giving crystallisation atreflux. The mixture was held at reflux for approximately 30 minutes andslowly cooled to ambient temperature. The mixture was further cooled andaged at 10-20° C. for 2 hours. The slurry was cooled to below 10° C. andthe product was filtered off, sucked dry and dried at approximately 60°C. under vacuum overnight to leave a white solid (4.34 g, 71%).

[0248] A more pure sample of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester unsolvated Form 2 was obtained by a coolingcrystallisation of 6α, 9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acidS-fluoromethyl ester (prepared eg according to Example 1, first method)in methanol (60 volumes, distilled at atmospheric pressure to approx37.5 volumes). The product was isolated by filtration and oven dried at60° C. under vacuum for 16 hours to leave a white, electrostatic solid(4.34 g, 71%).

[0249] The XRPD pattern of Example 2 product is shown in FIG. 1.

Example 3 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0250] Unsolvated Form 3

[0251] A suspension of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester acetone solvate (prepared eg according toExample 11) (20.0 g) in dichloromethane (800 ml, 40 volumes) and water(10 ml, 0.5 volumes) was heated to reflux giving a bright solution. Thesolution was clarified by hot filtration (Whatman 54 filter paper)during which time some solid crystallised in the filtrate which wasfully dissolved upon heating to reflux. The solution was distilled atatmospheric pressure (approx 400 ml solvent collected) and allowed tocool to ambient temperature. The mixture was further cooled and aged at<10° C. for 10 minutes. The product was filtered off, sucked dry anddried at approximately 60° C. under vacuum overnight to leave a whitesolid (12.7 g, 70%).

[0252] The XRPD pattern of Example 3 product is shown in FIG. 1 and FIG.4.

Example 4 Interconversion of Forms 1, 2 and 3 of Unsolvated 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0253] Slurrying a mixture of Form 1 and Form 2 in water at ambienttemperature revealed that the components are transformed entirely toForm 1 with time. XRPD results are shown in FIG. 2. Similar results wereobtained by slurrying a mixture of Form 1 and Form 2 in ethanol atambient temperature. From these results it may be concluded that Form 1is the thermodynamically more stable polymorphic form out of the twoforms.

[0254] Thermal XRPD studies on Form 3 were performed as shown in FIG. 4.The temperature and time profile is shown in FIG. 5 and the 5 tracesshown in FIG. 4 were obtained at the equilibration points shown in FIG.5. The results indicate that Form 3 is converted first to Form 2 andthen to Form 1 as temperature is elevated.

Example 5 Moisture Sorption of Forms 1, 2 and 3 of Unsolvated 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0255] The moisture sorption characteristics of the three forms weredetermined by monitoring the weight change of solid when exposed tostepwise increased and then decreased humidity. The results obtainedwere as follows:

[0256] Form 1: uptake of 0.18% w/w of moisture over the range 0-90%relative humidity at 25° C.

[0257] Form 2: uptake of 1.1-2.4% w/w of moisture over the range 0-90%relative humidity at 25° C.

[0258] Form 3: uptake of 1.2-2.5% w/w of moisture over the range 0-90%relative humidity at 25° C.

Example 6 Enthalpy of Dissolution of Forms 1 and 3 of Unsolvated 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0259] Enthalpies of dissolution in DMSO and acetonitrile weredetermined at 25° C. The results were as follows: Form 1 Form 3Acetonitrile +13.74 +8.62 DMSO  +1.46 −5.21

[0260] (results in kJ/mol)

[0261] Form these results it may be determined that the enthalpy oftransition from Form 3 to Form 1 is approximately 5.1-6.7 kJ/mol. On theassumption that the entropy of transition is small, since both Forms areunsolvated, the enthalpy of transition may be equated with the freeenergy of transition. Thus these data suggest that Form 1 is thethermodynamically most stable form at 25° C.

Example 7 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0262] Methylethylketone Solvate

[0263] A suspension of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (prepared eg according to Example 1) (400 mg)in methylethylketone (3.2 ml) is heated to reflux giving a clearsolution. A portion of the solvent is distilled off at atmosphericpressure (approx 1 ml) and the mixture cooled to approximately 20° C.The crystallised product is filtered off, dried at approximately 20° Cunder vacuum to leave the title compound as a white solid (310 mg, 68%).NMR δ (CDCl₃) includes the peaks described in Example 1 for the parentcompound and the following additional solvent peaks: 2.45 (2H, q), 2.14(3H, s), 1.06 (3H, t).

Example 8 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0264] Isopropanol Solvate

[0265] A solution of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (prepared eg according to Example 1) (150 mg)in isopropanol (15 ml) is left to slowly crystallise over a period ofapproximately 8 weeks. The resultant chunky crystals are isolated byfiltration to leave the title compound as a white solid. NMR δ (CDCl₃)includes the peaks described in Example 1 for the parent compound andthe following additional solvent peaks: 4.03 (1H, m), 1.20 (6H, d).

Example 9 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0266] Tetrahydrofuran Solvate

[0267] A suspension of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (prepared eg according to Example 1) (150 mg)in THF (20 vol) is warmed to give a clear solution. The solvent isallowed to slowly evaporate over a period of 6 days to leave titlecompound as a white solid. Alternatively, the THF solution is addeddropwise to solution of potassium bicarbonate (2% w/w) in water (50 vol)and the precipitated product collected by filtration to furnish thetitle compound as a white solid. NMR δ (CDCl₃) includes the peaksdescribed in Example 1 for the parent compound and the followingadditional solvent peaks: 3.74 (4H, m), 1.85 (4H, m).

Example 9 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0268] Tetrahydrofuran Solvate (Alternative Method)

[0269] A mobile suspension of 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid triethylamine salt (prepared eg according to Intermediate 1B) (1.2g) in THF (10 ml) is treated with a phase transfer catalyst(tetrabutylammonium bromide, typically between 8 and 14 mol %), cooledto approximately 3° C. and treated with bromofluoromethane (0.98equivalents). The suspension is stirred for between 2 and 5 hours,allowing to warm to 17° C. The reaction mixture is poured into water (30vol), stirred at approximately 10° C. for 30 minutes and filtered off.The collected solid is washed with water (4×3 vol) and the product ovendried under vacuum at 60° C. overnight to give the title compound as awhite solid (0.85 g, 87%).

Example 10 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0270] DMF Solvate

[0271] A mixture of Intermediate 1 (4.5 g, 8.88 mmol) in DMF (31 ml) istreated with potassium bicarbonate (0.89 g, 8.88 mmol) and the mixtureis cooled to −20° C. A solution of bromofluoromethane (0.95 g, 8.50mmol, 0.98 eqv.) in DMF (4.8 ml) at 0° C. is added and the mixture isstirred at −20° C. for 4 hours. The mixture is then stirred at −20° C.for a further 30 minutes, added to 2M hydrochloric acid (100 ml) andstirred for a further 30 minutes at 0-5° C. The precipitate collected byvacuum filtration, washed with water and dried at 50° C. to give thetitle compound (4.47 g, 82%). NMR δ (CD₃OD) includes the peaks describedin Example 1 for the parent compound and the following additionalsolvent peaks: 7.98 (1H, bs), 2.99 (3H, s), 2.86 (3H, s).

Example 11 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester

[0272] Acetone Solvate

[0273] A solution of Intermediate 1 (530.1 g, 1 wt) in dimethylformamide(DMF) (8 vol) is treated with potassium hydrogen carbonate (0.202 wt,1.02 eq) and the mixture cooled to −17±3° C. with stirring.Bromofluoromethane (BFM) (0.22 wt, 0.99 eq) is then added and thereaction stirred at −17±3° C. for at least 2 h. The reaction mixture isthen added to water (17 vol) at 5±3° C. over ca 10 min followed by awater (1 vol) line wash. The suspension is stirred at 5-10° C. for atleast 30 min and then filtered. The filter cake (the DMF solvate of 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester) is washed with water (4×4 vol) and theproduct is pulled dry on the filter. The damp cake is returned to thevessel, acetone (5.75 vol) added and heated at reflux for 2 h. Themixture is cooled to 52±3° C. and water (5.75 vol) added, keepingtemperature at 52±3° C. The mixture is then cooled to 20±3° C., filteredand dried in vacuo at 60±5° C. overnight to give the title compound as awhite solid (556.5 g, 89%). NMR δ (CDCl₃) includes the peaks describedin Example 1 for the parent compound and the following additionalsolvent peaks: 2.17 (6H, s).

Example 12 Dry Powder Composition Containing 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester, Unsolvated Form 1

[0274] A dry powder formulation was prepared as follows: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy- 0.20 mg16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid S-fluoromethylester, unsolvated Form 1 (prepared according to Example 1, firstalternative method and micronised to a MMD of 3 μm): milled lactose(wherein not greater than 85% of particles 12 mg have a MMD of 60-90 μm,and not less than 15% of particles have a MMD of less than 15 μm):

[0275] A peelable blister strip containing 60 blisters each filled witha formulation as just described was prepared.

Example 12A Dry Powder Composition Containing 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester, Unsolvated Form 1, and a Long Actingβ₂-adrenoreceptor agonist

[0276] A dry powder formulation may be prepared as follows: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy- 0.20 mg16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid S-fluoromethylester, unsolvated Form 1 (prepared according to Example 1, firstalternative method and micronised to a MMD of 3 μm): Long-actingβ₂-adrenoreceptor agonist (micronised to a MMD 0.02 mg of 3 μm): milledlactose (wherein not greater than 85% of particles 12 mg have a MMD of60-90 μm, and not less than 15% of particles have a MMD of less than 15μm):

[0277] A peelable blister strip containing 60 blisters each filled witha formulation as just described may be prepared.

Example 13 Aerosol Formulation Containing 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester, Unsolvated Form 1

[0278] An aluminium canister was filled with a formulation as follows:6α, 9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy- 250 μg16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid S-fluoromethylester, Unsolvated Form 1 (prepared according to Example 1, firstalternative method) and micronised to a MMD of 3 μm):1,1,1,2-tetrafluoroethane: to 50 μl

[0279] (amounts per actuation) in a total amount suitable for 120actuations and the canister was fitted with a metering valve adapted todispense 50 μl per actuation.

Example 13A Aerosol Formulation Containing 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester, Unsolvated Form 1, and a Long Actingβ₂-adrenoreceptor agonist

[0280] An aluminium canister may be filled with a formulation asfollows: 6α, 9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy- 250μg 16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acidS-fluoromethyl ester, Unsolvated Form 1 (prepared according to Example1, first alternative method) and micronised to a MMD of 3 μm):Long-acting β₂-adrenoreceptor agonist (micronised to a MMD 25 μg of 3μm): 1,1,1,2-tetrafluoroethane: to 50 μl

[0281] (amounts per actuation) in a total amount suitable for 120actuations and the canister may be fitted with a metering valve adaptedto dispense 50 μl per actuation.

Example 14 Nasal Formulation Containing 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester, Unsolvated Form 1

[0282] A formulation for intranasal delivery was prepared as follows:6α, 9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy- 10 mg16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid S-fluoromethylester Unsolvated Form 1 (prepared according to Example 1, firstalternative method, micronised): Polysorbate 20 0.8 mg Sorbitanmonolaurate 0.09 mg Sodium dihydrogen phosphate dihydrate 94 mg Dibasicsodium phosphate anhydrous 17.5 mg Sodium chloride 48 mg Demineralisedwater to 10 ml

[0283] The formulation was fitted into a spraypump capable of deliveringa plurality of metered doses (Valois).

[0284] Throughout the specification and the claims which follow, unlessthe context requires otherwise, the word ‘comprise’, and variations suchas ‘comprises’ and ‘comprising’, will be understood to imply theinclusion of a stated integer or step or group of integers but not tothe exclusion of any other integer or step or group of integers orsteps.

[0285] The patents and patent applications described in this applicationare herein incorporated by reference.

1. A pharmaceutical formulation for administration by inhalationcomprising a compound of formula (I),

or a solvate thereof, together with a long-acting β₂-adrenoreceptoragonist which formulation has a therapeutically useful effect in thetreatment of inflammatory disorders of the respiratory tract over aperiod of 24 hours or more.
 2. A pharmaceutical formulation according toclaim 1 wherein the compound of formula (I) or a solvate thereof and thelong-acting β₂-adrenoreceptor agonist are both present in particulateform.
 3. A pharmaceutical formulation according to claim 2 furthercomprising a particulate carrier.
 4. A pharmaceutical formulationaccording to claim 3 wherein the carrier is lactose.
 5. A pharmaceuticalformulation according to claim 1 further comprising a liquifiedpropellant gas.
 6. A pharmaceutical formulation according to claim 2further comprising a liquified propellant gas.
 7. A pharmaceuticalformulation according to claim 1 wherein the inflammatory disorder ofthe respiratory tract is asthma.
 8. A method of treatment of ainflammatory disorder of the respiratory tract once-per-day whichcomprises administration of a pharmaceutical formulation according toclaim
 1. 9. A method of treatment according to claim 8 wherein theinflammatory disorder of the respiratory tract is asthma.
 10. An inhalercontaining a plurality of doses of a pharmaceutical formulationcomprising a compound of formula (I)

or a solvate thereof, together with a long-acting β₂-adrenoreceptoragonist, which formulation has a therapeutically useful effect in thetreatment of inflammatory disorders of the respiratory tract over aperiod of 24 hours or more, and which doses are suitable foronce-per-day administration of the formulation by inhalation.
 11. Aninhaler according to claim 10 wherein the compound of formula (I) or asolvate thereof and the long-acting β₂-adrenoreceptor agonist are bothpresent in particulate form.
 12. An inhaler according to claim 11wherein the formulation further comprises a particulate carrier.
 13. Aninhaler according to claim 12 wherein the carrier is lactose.
 14. Aninhaler according to claim 10 wherein the formulation further comprisesa liquefied propellant gas.
 15. An inhaler according to claim 11 whereinthe formulation further comprises a liquefied propellant gas.
 16. Aninhaler containing a plurality of doses of a pharmaceutical formulationcomprising a particulate compound of formula (I)

or a solvate thereof, a particulate long-acting β₂-adrenoreceptoragonist and a carrier, each drug being present in an amount adequate toprovide a therapeutically useful effect in the treatment of inflammatorydisorders of the respiratory tract over a period of 24 hours or morefollowing once-per-day dosing by inhalation.
 17. An inhaler according toclaim 10 wherein wherein the inflammatory disorder of the respiratorytract is asthma.
 18. An inhaler according to claim 16 wherein whereinthe inflammatory disorder of the respiratory tract is asthma.